In healthy conditions, assimilation, storage and utilization of nutrient energy constitute a highly integrated homeostatic system. Maintaining a relatively constant level of energy stores, and hence body weight, requires the achievement of a balance between food intake and energy expenditure. The “set point” hypothesis proposes coordinated regulation by control centers in the central nervous system. Hypothalamic nuclei are believed to be the sites at which the “set point” is regulated, given their important role in establishing homeostasis through regulation of food intake (hunger versus satiety), body weight, energy expenditure (adaptive thermogenesis) and hormone integration involving substrate inter-conversion, storage and mobilization as appropriate.
An expanding array of (neuro) peptides and neurotransmitters are being discovered that alter food intake when administered peripherally or directly into the hypothalamus. Mouse models of obesity include five apparently single gene mutations. The most intensively studied mutations occur in the ob and db genes which are intimately involved in the control of body fat deposits. This control involves a signaling pathway including a hormone secreted by adipocytes and acting through specific receptors in areas of the brain that govern ingestive behavior and metabolic activity (Flier, 1997; Flier and Maratos-Flier, 1998; Spiegelman and Flier, 1996).
Cloning of the mouse ob gene and its human homologue led to the identification of the ob gene product (ob protein) now named leptin (Zhang et al., 1994). It is secreted primarily by white adipocyte tissue as a non-glycosylated 146-amino acid polypeptide with a MW of 16 kDa. Consistent with its role as the postulated adipose-derived satiety factor, administration of recombinant leptin to ob/ob mice caused rapid reversal of the obese phenotype (Campfield et al., 1995; Halaas et al., 1995; Pelleymounter et al., 1995) by decreasing food intake and increasing energy expenditure. Furthermore, administration of leptin corrects most, if not all of the metabolic and endocrine defects, including sterility, in the ob/ob mice (Chehab et al., 1996).
Leptin displays no apparent sequence similarity to any other known protein. However, based on structure prediction analysis, leptin is identified as a member of the hematopoietic cytokine receptor family carrying the typical four aa-helical bundle structure. Accordingly, its receptor belongs to the class 1 cytokine receptor superfamily. As a cytokine, leptin has a pleiotropic role including effects on the hemopoietic system, immune defense and inflammation, reproduction and pregnancy, regulation of renal function and acceleration of the onset of puberty in female rodents. In addition to its central hypothalamic action, leptin may also exert effects on extraneural tissues extending its metabolic effects. This extended metabolic effects include, repression of fatty acid and lipid synthesis, triacylglyceride depletion of tissues and increased expression of enzymes of fatty acid metabolism and of the uncoupling protein UCP-2, believed to play an important role in thermogenesis. Recent studies show that leptin also plays a role in hematopoiesis, fatty acid homeostasis in cells, hepatic metabolism and protection against TNF-induced toxicity. Leptin also stimulates the proliferation of CD4+T-cells and has a direct effect on endothelial cells, and cells of the gastro-intestinal tract.
Leptin exerts its effects upon binding to a high affinity receptor, the product of the db gene. This receptor was originally cloned from mouse choroid plexus and may exist in at least six isoforms through alternative splicing (Tartaglia et al.,1995; Lee et al.,1996). So far, most attention has been focused on a ubiquitously expressed splice variant with short cytoplasmic tail (OB—Ra), and on the isoform with a long cytoplasmic domain (OB—Rb), which is predominantly expressed in certain nuclei in the hypothalamus. RT-PCR and Northern blot analysis also showed expression of the latter isoform in peripheral tissues including pancreas, liver, lung, kidney and adrenals, intestine and adipose tissues, as well as in endothelial cells and CD4+helper T-cells.
The detailed structure of the murine leptin receptor gene was recently described. In the db/db mice, a point mutation causing the use of a cryptic splice site leads to the expression of a receptor with a truncated cytoplasmic tail which is likely signaling deficient. The fatty (foreign associate) gene in rats appears to be a functional homologue of the mouse db gene, due to a Gln269Pro substitution in the extracellular domain of the receptor, leading to impaired signaling and obesity. Underscoring the evolutionary conserved role of this pathway, homologous mutations were recently described in humans. Severe early-onset obesity was observed in patients with mutations leading to either impaired leptin (Montague et al., 1997) or leptin receptor (Clement et al., 1998) function.
Based on sequence homology and functional aspects, the leptin receptor is considered a member of the Class I cytokine receptor superfamily, and is most closely related to gp 130, the signaling component of the IL-6R complex, and to the LIF and G-CSF receptors. As such, the leptin receptor contains typical motifs involved in signaling such as a JAK tyrosine kinase binding site and a Box 3 involved in recruitment of STAT-3 upon ligand-induced receptor tyrosine phosphorylation. The long isoform is generally believed to be the signaling competent receptor although divergent signaling capacities have been described for the long and short isoforms which both contain the Box 1 motif. The receptor functions as a (ligand-induced) homodimer, independent of gp 130. Leptin binding leads to the activation of JAK2 and multiple STATs (STAT-1, STAT-3 and STAT-5b), however only STAT-3 activation was observed in hypothalamic centers in vivo. In established hepatoma cell lines stably expressing the OB—R long isoform, the leptin receptor appears to be functionally equivalent to the endogenous IL-6R.
One target for leptin action in the hypothalamus is neuropeptide Y (NPY), a key effector of nutritional homeostasis that stimulates appetite. Leptin induces inhibition of NPY biosynthesis and release. The observation that NPY-deficient mice did not completely reverse the obesity phenotype, although NPY is required for full manifestation of the ob/ob phenotype, makes it likely that other leptin targets must exist. Such alternative target candidates include glucagon-like peptide 1 (GLP-1) produced in the brain stem and causing reduced food intake, the melanin-concentrating hormone also involved in hypothalamic regulation of ingestive behavior, the hypothalamic corticotropin-releasing factor (CRF) inhibiting appetite and stimulating metabolism, and the recently described orexins and cocaine- and amphetamine-regulated transcript (CART), a hypothalamic satiety factor. Effects on the expression of the pro-opio-melanocortin gene by leptin have also been described.
Obese humans, except patients carrying the rare, previously identified, mutations in the genes for leptin or its receptor, generally produce higher levels of circulating leptin, suggesting that obesity is associated with “leptin resistance”. Such resistance could conceivably occur at several levels: peripheral leptin dysfunction, dysregulation of the saturable leptin transport through the blood brain barrier, and the expression of, and signaling by, the hypothalamic leptin receptor. Variable mechanisms leading to leptin resistance are suggested by studies in murine models (Halaas et al., 1997).